Currently, an expression system using Escherichia coli as a host is widely used for preparing a large amount of a recombinant protein derived from a eukaryote. This is because the system is readily handled and its research progresses most (Weickert et al., Curr. Opin. Biotechnol. 7: 494-499 [1996]).
On the other hand, the present inventors have previously demonstrated that Rhodococcus erythropolis can also be used as a host for recombinant protein production (JP Patent Publication (Kokai) No. 2004-73032A and Japanese Patent Application No. 2002-235008). R. erythropolis is one of Actinobacteria capable of proliferation at temperatures ranging from 4° C. to 35° C. The biggest feature of an expression system that uses this bacterium as a host is in that a recombinant protein can be produced at temperatures not higher than 10° C. such as 4° C. In systems that employ other hosts including Escherichia coli, bacteria belonging to the genus Bacillus, yeasts, and Sf9 insect cells (Cereghino and Cregg, Curr. Opin. Biotechnol. 10 422-427 [1999]; and Miller, Curr. Opin. Genet. Dev. 3 97-101 [1993]), recombinant protein production at temperatures not higher than 10° C. is quite difficult. Recombinant protein production at temperatures not higher than 10° C. has allowed the production of proteins difficult to produce until then, for example, proteins inhibiting the proliferation of host cells, proteins getting insoluble at temperatures around 30° C., and proteins derived from organisms adapting to low temperatures.
The present inventors have constructed a group of expression vectors for a bacterium belonging to the genus Rhodococcus called pTip vectors and have used them in recombinant protein production (JP Patent Publication (Kokai) No. 2004-73032A and Japanese Patent Application No. 2002-235008). These vectors contain the promoter of a TipA gene whose expression is induced by antibiotic thiostrepton and also contain a multiple-cloning site (MCS) for cloning a foreign gene (gene to be expressed), located downstream of the promoter. Thus, the pTip vectors are thiostrepton-inducible expression vectors. In bacteria belonging to the genus Rhodococcus that are transformed with these expression vectors, foreign protein production is induced only when thiostrepton is added to a culture solution.
Non-Patent Document 1
Weickert et al., Curr. Opin. Biotechnol. 7: 494-499 [1996]
Non-Patent Document 2
Cereghino and Cregg, Curr. Opin. Biotechnol. 10 422-427 [1999]
Non-Patent Document 3
Miller, Curr. Opin. Genet. Dev. 3 97-101 [1993]